Directed enzyme evolution via small and effective neutral drift libraries.
Small libraries for directed evolution can be obtained by neutral drifts that maintain the protein's original function, yielding highly polymorphic, stable and evolvable variants. We describe methods for preparing such libraries, using serum paraoxonase (PON1). An optimized GFP variant fused to PON1 reported levels of soluble, functional enzyme, enabling selection by flow cytometry and identification of enzyme variants exhibiting improved specific and total activities toward several substrates, including toxic organophosphates.
Gupta
Rinkoo D
RD
Department of Biological Chemistry, Weizmann Institute of Science, Rehovot 76100, Israel.
Tawfik
Dan S
DS
D043303:Aryldialkylphosphatase / Q000032:analysis* / Q000235:genetics / Q000378:metabolism*; D019020:Directed Molecular Evolution; D017930:Genes, Reporter / Q000235:genetics; D040961:Genetic Drift; D009154:Mutation / Q000235:genetics; D019151:Peptide Library
D019151:Peptide Library; D043303:Aryldialkylphosphatase